Construction of a Subtractive cDNA Library from the Paralichthys olivaceus Embryonic Cells Induced by a Double-Stranded RNA Virus
Abstract: Using suppression subtractive hybridization (SSH) technique, a subtractive cDNA library was constructed from Japanese flounder (Paralichthys olivaceus) embryonic cells (FEC) induced by UV-inactivated dsRNA virus GCHV (Grass carp hemorrhage virus). A housekeeping gene, α-tublin, was used to estimate the efficiency of subtractive cDNA. In this library, α-tublin was subtracted at about 2~(10) folds, indicating that some differentially expressed genes were also enriched at about the same folds. The length of the subtractive cDNA fragments cloned into pGEM-T vector ranged from 250bp to 2000bp.The results showed that the subtractive cDNA library is successful, which will be very useful for the understanding of antiviral immune response to dsRNA virus and essential for rapid isolation of differentially expressed genes induced by dsRNA virus.